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teritis in piglets associated with Aujeszky’s disease virus infection. Vet Pathol 21:450-452. 14. Patel JR, Edington N, Mumford JA: 1982, Variation in cellular tropism between isolates of equine herpesvirus-1 in foals. Arch Virol 74:41-51. 15. Sambrook J, Fritsch EF, Maniatis T: 1989, Molecular cloning: a laboratory manual, 2nd ed. Cold Spring Harbor Laboratory’ Press, Cold Spring Harbor, NY. 16. Studdert MJ: 1983, Restriction endonuclease DNA fingerprinting of respiratory, foetal and perinatal foal isolates of equine herpesvirus type 1. Arch Virol 77:249-258. 17. Studdert MJ, Simpson T, Roizman B: 1981, Differentiation of respiratory and abortigenic isolates of equine herpesvirus 1 by restriction endonucleases. Science 214:562-564. 18. Whitwell KE, Blunden AS: 1992, Pathological findings in horses dying during an outbreak of the paralytic form of equid herpesvirus type 1 (EHV1) infection. Equine Vet J 24: 13-19. 19. Yeargan MR, Allen GP, Bryans JT: 1985, Rapid subtyping of equine herpesvirus 1 with monoclonal antibodies. J Clin Microbiol 21:694-697.

Leber hereditary optic neuropathy (LHON) is a maternally inherited disease characterized by acute bilateral loss of central vision.1 In approximately 95% of the cases worldwide, LHON is caused by 3 pathogenic point mutations in mitochondrial DNA (mtDNA) coding for the respiratory chain subunits of complex I genes (the reduced form of nicotinamide adenine dinucleotide–ubiquinone-oxidoreductase), namely m.3460G>A in ND1, m.11778G>A in ND4, and m.14484T>C in ND6.2 Of these 3 mutations, 11778G>A, resulting in an arginine to histidine substitution at amino acid 340, is responsible for half of all LHON cases. Patients with this mutation exhibit the poorest prognosis for spontaneous visual improvement, and there is no effective therapy.3 Most patients with LHON carry the mtDNA mutations in homoplasmic condition, that is, they have no normal ND4 mtDNA. Thus, an effective therapeutic approach would be introduction of normal mtDNA into the affected cells (ganglion cells of the retina) in these patients. One of the major limitations in this aspect is that few practical methods for delivering genes to the mitochondria are available.4,5 To address this, we and other groups adapted an approach termed allotopic expression, in which the mutant mitochondrial gene (in this case ND4) is expressed in the nucleus, translated on cytoplasmic polyribosomes, and transported into mitochondria with the addition of an N-terminal mitochondrial targeting sequence.6-8 Allotopic expression of the normal ND4 gene normalized the defective adenosine triphosphate (ATP) synthesis of LHON cells and improved cell survival.9 Recombinant adeno-associated virus (AAV) vectors have been used safely for ocular gene delivery in a disorder affecting photoreceptors, Leber congenital amaurosis.10,11 Adeno-associated virus is derived from the nonpathogenic virus belonging to the Dependovirus genus of the Parvoviridae family and contains a linear single-stranded DNA genome enclosed in a capsid composed of 3 proteins: VP1, VP2, and VP3.12,13 Studies on the hybrid vectors or the pseudotyped vectors containing the promoter and the trans-gene flanked by the 145–base pair (bp) AAV inverted terminal repeats packaged into different capsid serotypes not only reveal cellular tropism in different organs but also show an increased efficiency for specific tissues.14,15 In a previous study, we showed that the human ND4 subunit was expressed in only one-third of retinal ganglion cells (RGCs) 1 month after intravitreal injections in mice. For treatment of acute visual loss in LHON, a more efficient and rapid gene expression delivery system is needed. Strategies to improve the efficiency of transgene expression are being continuously explored by manipulating the AAV genome. One such development is the self-complementary AAV (scAAV) vector with a double-stranded vector genome. This vector was generated by deleting the terminal resolution site from one of the inverted terminal repeats of recombinant AAV (rAAV), which prevents the initiation of replication at the mutated end.16 This strategy allows packaging of a self-complementary form of vector DNA that has the ability of annealing to itself to form a double-stranded DNA immediately on vector delivery to the target cell nucleus. This overcomes the rate-limiting step of replication of the single-stranded viral genome into the double-stranded DNA within the cell nucleus, thereby increasing the efficiency and speed of transgene expression manyfold, as observed in the liver,17,18 brain,19 eye,20,21 and heart.22 Whether scDNA is single or double stranded while inside the AAV vector capsid is unclear at present. In this study, we compared expression of the human ND4 subunit gene when delivered by scAAV relative to that when delivered by the single-stranded AAV (ssAAV).

SUMMARYComplement and antibody contribute to infection-enhancement and possible expanded cellular tropism of HIV-1 in vitro through a process requiring complement receptors. Until now, however, the ability of HIV-1 to bind complement receptors has not been documented or characterized. We investigated whether antibody and complement permitted HIV-1 to bind to the B lymphocyte receptor, CR2 (CD21), in an effort to learn more about infection-enhancement, and also because CR2 can mediate B cell proliferation and antigen localization in lymphoid organs in other systems. HIV-1 incubated with antibody and fresh human serum as a source of complement bound approximately 10-fold greater to cells expressing CR2 than to HIV-1-permissive cells lacking this receptor. A similar effect was observed using cells which expressed CR2 but no CD4. This binding was minimal in heat-inactivated and C3-deficient sera, and was significantly reduced by the anti-CR2 MoAb, OKB7, but not by the anti-CD4 MoAb, OKT4a. Thus, complement and antibody acted in concert to facilitate the binding of HIV-1 to CR2 independently of CD4. CD4-independent binding of HIV-1 to CR2 was not sufficient to produce infection in Raji-3 cells. Titres of antibodies mediating CR2 binding correlated with antibody titres as measured by immunofluorescenee (P < 0.01) and infection-enhancement (P < 0.05) but were discordant with titres of neutralizing antibodies, a result consistent with the utilization of CR2 for enhanced infection of cells. The ability of complement and antibody to facilitate the binding of HIV-1 to CR2 in the absence of CD4 provides new insights into mechanisms of HIV-1-induced immunopathogenesis and infection-enhancement.

We investigated the infection of peripheral blood mononuclear cells (PBMNC) by hepatitis C virus (HCV) in 5 patients with HCV-related chronic hepatitis. The presence of HCV-RNA-positive and -negative strands was tested with the polymerase chain reaction (PCR) method. In all subjects, HCV-RNA was shown in PBMNC. In 3 cases, HCV-RNA was shown in the T- and B-cell populations, with viral RNA also present in the monocyte-macrophage fraction of two of these. HCV-RNA-negative stranded molecules, indicative of the viral multiplication, were significantly increased in cells maintained in cultures with PHA/PMA stimulation. The results indicate that HCV infect blood mononuclear cells, thus suggesting that this cellular tropism may play a role in HCV infection.

An assay based on production of HIV antigen in cultures of CD4+ lymphocytes infected ‘in vitro’ with cell-free virus was established. Using this assay it was possible to isolate, propagate and reliably determine the zidovudine susceptibility of HIV isolates from all patients despite differences in cellular tropism and syncytium inducing capacity. Using this assay, differences in zidovudine susceptibility of 52 serial isolates obtained from 16 patients before and after initiation of therapy were examined. HIV with a 10- to 100-fold reduced susceptibility to zidovudine were isolated from 13 patients as early as 4 months after initiation of therapy. Number of months of zidovudine treatment was strongly associated with development of viral resistance, and high CD4 cell counts tended to be associated with lower rates of development of resistance. That patients can harbor mixtures of virus strains with different susceptibility to zidovudine was confirmed by the differences in susceptibility between isolates obtained simultaneously from CD4+ lymphocyte and plasma, and by the differences in susceptibility between virus strains isolated from clones of CD4+ lymphocytes.

Publisher Summary Exanthem subitum or roseola infantum is a common disease in infants, characterized by high fever for a few days and the appearance of rash coinciding with subsidence of fever. It has been believed that a virus causes this disease because of evidences of the long incubation time, the leukopenia, the failure to respond to antimicrobial agents, and the failure to recover bacteria from the various body fluids. However, attempts to isolate the causative agents have been unsuccessful. This chapter illustrates that HHV-6 is the causal agent of exanthema subitum. This chapter also gives a walkthrough that HHV-6 causes exanthem subitum and this virus may have a cellular tropism of CD4+ lymphocyte. The airborne route may be suggested as a mode of transmission of HHV-6 infection.

Although human immunodeficiency virus (HIV) is the causative agent of the acquired immunodeficiency syndrome and related disorders, it has been suggested that viral cofactors may accelerate the progression of the disease. We present evidence that human T lymphoid cells productively coinfected by HIV type 1 (HIV-1) and human T-cell leukemia virus type I (HTLV-I) or HTLV-II generate a progeny of phenotypically mixed viral particles that allow the penetration of HIV-1 into previously nonsusceptible CD4- human cells, including mature CD8+ T lymphocytes, B lymphoid cells, epithelial cells, and skeletal muscle cells. The infection is independent of the major HIV-1 receptor, (i.e., the CD4 glycoprotein) since OKT4a, a neutralizing anti-CD4 monoclonal antibody, fails to block the penetration of HIV-1. Similarly, infection is not inhibited by monoclonal antibody M77, directed toward the neutralizing loop of the gp120 envelope glycoprotein of HIV-1. In contrast, pretreatment of the virus stock with HTLV-I-neutralizing human serum completely abolishes the penetration of phenotypically mixed HIV-1 into CD4- cells. These results suggest that HTLV-I or HTLV-II may increase the pathogenicity of HIV-1 by broadening the spectrum of its cellular tropism and, thus, favoring its spread within the organism of coinfected hosts.

The discovery that members of the chemokine receptor family act as necessary co-receptors for entry of human immunodeficiency virus-I (HIV-1) into susceptible cells has provided major insights into the determinants of celLular tropism. Variations in the ability of certain strains to replicate in different CD4 + cells can now be explained by unique interactions between the viral envelope and any one of a number of co-receptors expressed on different CD4+ cells. Viral strains previously characterized as predominantly T-tropic and with the capability of replicating in tumor derived T-cell lines utilize the CXCR4 receptor, while macrophage-tropic isolates primarily use CCR-5 [1]. At least nine additional chemokine receptor-like molecules have now been demonstrated to be capable of acting as co-receptors for HIV using in vitro assays; however, the role of many of these additional coreceptors in vivo remains unclear, as the majority of primary isolates have been shown to use CXCR4, CCRS and CCR3 alone or in combination (21. Tissue specific selective expression of these co-receptors most likely plays a role in determining tissue tropism in vwo, thus potentially influencing different disease manifestations. Furthermore, genetic polymorphism of either the chemokine receptors or their ligands has provided insight into the genetic determinants of infection as well as disease progression. Individuals homozygous for a deletion in one of the coreceptors, CCR5, are rarely infected with HEy-i. Interestingly, this deletion is predominantly found in those of Northern European descent [3]. HIV-infected people displaying a slow disease progression have been found to have one or more genetic polymorphisms including heterozygous deletion of CCR5, polymorphism in an alternative coreceptor CCR2b or in the CXCR4 receptor ligand, SDF-1 (4, 51 HIV-associated nephropathy (HIVAN) is a unique tissue manifestation of infection with a striking predominance in African Americans [6]. A full evaluation of the renal expression of HIV-1 co-receptors as well as viral strain utilization of these coreceptors in individuals with this manifestation might shed light on both the pathogenesis as well as genetic determinants of HIVAN. HIVAN, like HIV-associated central nervous system (CNS) disease, is a relatively late complication, develops in only a subset of individuals infected with the virus, and to date appears to be a direct complication of the virus, although the role of a cofactor(s) cannot be excluded. The direct demonstration of the virus has been more convincingly demonstrated in primary CNS cells, particularly microglial cells. Direct infection of primary renal cells in vitro and in viva has been problematic to date, although it is always difficult to exclude technical difficulties as well as the limitations of in vitro systems. In studies directed at determining the role of coreceptor specificity in CNS disease, several of these coreceptors have been shown to facilitate infection of cells derived from the CNS, although there is not a full understanding of how this selective expression determines the development of HI V-associated CNS complications in certain individuals. CCR5 and possibly CCR3 can act as coreceptors in the presence of CD4 for microglia, one of the major targets for CNS infection. Interestingly, some of the more recently described coreceptors, including STRL33 and TERI/CCR8, are expressed in brain-derived cells; however, their specific role in determining infection of the central nervous system remains unclear [7]. The SIV/macaque model has provided some interesting insights into the potential role of these coreceptors in CNS disease that may have implications for HIV-1. Brain capillary endothelial cells from rhesus macaques express the co-receptors CCRS, CCR3, CXCR4 and STRL33, and can be infected by a particularly neurovirulent strain of Sly. Furthermore, entry into these endothehal cells using the co-receptor CCR5 appears to be uniquely CD4-independent [8]. The issue of tissue-specific expression of HEy-I coreceptors as one of the selective events in HIVAN is appropriate to consider as well. The importance of this issue does not necessarily depend on the direct demonstration of the virus in the kidney, as a circulating viral envelope would be sufficient to bind to and induce signaling through any one of these seven-transmembrane-spanning G-protein coupled receptors. As a step in exploring a role for these co-receptors in HI VAN, the article by Eitner et al in this issue of Kidney International evaluates the expression of CCR5 in human and primate kidneys in the presence and absence of HIV or Sly infection [9]. Using in situ hybridization, the authors did not detect CCR5-specific rnRNA in intrinsic renal cells of normal kidneys nor in kidneys from HIV-1, HIV-2 or SIV-infected macaques. None of the kidneys from the lentivirus-infected animals had pathologic features consistent with 1-H VAN. However, in human kidneys exhibiting a significant inflammatory reaction as seen in interstitial nephritis and/or associated with transplant rejection, CCR5 expression was detected and appeared to be localized to either infiltrating mononuclear cells and/or endothelial or vascular smooth muscle cells. The absence of expression of this co-receptor in kidneys with no significant inflammatory process does not exclude the role of this co-receptor in HIVAN, either through direct infection of intrinsic renal cells or indirectly through tissue infiltration with infected cells or viral products. Expression of CCR5 in mononuclear-derived cells is regulated by a number of stimuli including activation and differentiation (10]. If expression of CCR5 on primary renal cells was involved in HIVAN, it is possible that cytokine stimulation is necessary as a prerequisite event, and therefore examination of kidneys with early pathologic features of HIVAN might provide further insight into the role of this co-receptor. More intriguing is the possibility that an alternative co-receptor, perhaps one that is infrequently expressed or for which there is considerable genetic polymorphism, is involved in either entry of certain HIV-1 strains into renal tissue or binding to circulating envelope. This would provide a potential explanation for the striking predominance of this disease in blacks. The pathogenesis of HIVAN remains an enigma. New leads provided by the discovery of HIV co-receptors should be vigorously pursued and may not only provide insights into pathogenesis, but provide new approaches to therapy as well. A complete characterization of co-receptor expression in primary intrinsic renal cells under normal and pathologic conditions, as well as further characterization of viruses and viral envelopes derived from renal tissue of individuals with this devastating manifestation, will be necessary steps in this process.

In the last 15 years there have been numerous reports demonstrating that transplanted hepatocytes can retain hepatocellular functions (1), correct congenital enzyme deficiencies (2–4), and improve survival rates in animal models of acute hepatic failure (5–7). Recently, Moscioni et al. (8) showed that human liver cells attached to collagen-coated microcarriers (9, 10) were able to correct metabolic defects following transplantation into mutant rat recipients that were genetically deficient in either bilirubin-uridine diphosphate glucuronosyltransferase activity or albumin synthesis. Interestingly, the intraperitoneally injected hepatocyte-microcarriers that were well vascularized were almost exclusively found on the surface of the pancreas. This apparent cellular tropism raises questions about the presence and mechanism of action of pancreas-derived hepatotrophic factors and their role in the maintenance of hepatocyte integrity (11–14). The aim of this study was to determine the effect of pancreatic islets on the survival of human hepatocytes in an ectopic site in nude mice. Human hepatocytes were obtained from segments of preserved cadaver donor livers that had been discarded during size reduction procedures before transplantation of the residual fragments to pediatric recipients. A modification of the automated procedure for isolation of human pancreatic islets (15) was used for the isolation of hepatocyte aggregates. After cannulation of its portal branch, the liver segment was perfused with 200 ml of a solution of 1 mg/ml collagenase (Boehringer-Mannheim, type P) in Hanks solution (28°C) and placed into a digestion chamber. Separation of hepatocellular clusters was accomplished by the same separation procedure used previously for isolation of human pancreatic islets (15). Between 80% and 95% of the harvested hepatocytes were viable, as assessed by trypan blue exclusion. Collagen-coated microcarriers (Cytodex 3, Pharmacia Fine Chemicals, Upsala, Sweden) were prepared as described by Demetriou et al. (9). Pancreatic islets were obtained from the pancreata of mongrel dogs (18–25 kg) by a modification (16) of the automated method for human islet isolation (15). Male BALB/c nude mice (20 g) were used as recipients in three groups (n=6 in each group). In group 1 (control) approximately 4000 hepatocyte clusters of an average diameter of 150 µ were placed beneath the left renal capsule as previously described (17). In group 2 (secondary control), hepatocyte clusters were transplanted in combination with 400 collagen-coated microcarriers. In group 3 the same number of hepatocyte clusters was transplanted without microcarriers but in combination with 400 pancreatic islets of 150 µm average diameter. Thirty days after transplantation the nude mice were sacrificed and the morphologic integrity of the renal subcapsular grafts was determined histopathologically. Hematoxylin-eosin stain was used in all sections. In addition, immunostain for insulin (immunoperoxidase) was used in sections of combined islet-hepatocyte transplants to distinguish the islets from the hepatocellular implants. The pathologist, who was blinded to the experimental groups from which the specimens came, correctly and without exception matched the tissues to the groups as follows. No hepatocytes were detectable in the renal subcapsular implants 1 month after transplantation of hepatocyte clusters alone (group 1). In group 2, the addition of collagen-coated microcarriers to the hepatocellular aggregates resulted in the survival of a thin rim of epithelioid cells attached to the microcarriers, while the remaining hepatocytes degenerated, leaving only fibrosis and a nonspecific mononuclear cell infiltrate around the microcarriers in the transplant site (Fig. 1). This suggested a minor augmentation of hepatocellular survival by the microcarrier. In contrast, when islets were included at the transplant site (group 3) multilayers of human hepatocytes were found. The surviving hepatocytes were healthy, in large clumps, and glycogen-containing, PAS-positive, and diastase-sensitive. These results are consistent with previous demonstrations that pancreatic-derived hepatotrophic factors are essential for maintenance of hepatocyte integrity (11–14). Scattered pancreatic islets were detected (Fig. 2). FIGURE 1 Photomicrograph of combined human hepatocytes and collagen-coated microcarriers 4 weeks after renal subcapsular transplantation. Degeneration of the hepatocytes occurred, leaving only a thin rim of epithelioid cells attached to the microcarriers (M) (H&E ... FIGURE 2 Photomicrograph of combined human hepatocytes and dog pancreatic islets 4 weeks after renal subcapsular transplantation in nude mice. Insulin immunoperoxidase (dark) stain distinguished islets (I) from the human hepatocytes (H), which appeared well preserved ... In past experience, when hepatocytes were transplanted by themselves in the presence of an intact host liver, no evidence of hepatocyte proliferation could be observed at any time after transplantation (18). Whenever cells without nuclei and multinucleated cells were found at the transplant site (19), hepatic degeneration had occurred. Similarly, it was not possible to detect the hepatocytes at either 2 or 4 days after intrasplenic transplantation (20), except when there was a proliferative stimulus provided by 70% recipient hepatectomy. Using the model of Demetriou et al. (9, 10), the collagen-coated hepatocyte microcarriers, the microcarriers may have promoted the survival of a small number of hepatocytes. In the earlier experiments of Moscioni et al. (8), the selective survival of the intraperitoneally injected hepatocyte-microcarrier aggregates on the surface of the pancreas was congruent with the possible role of factors produced from the pancreas in the maintenance of normal hepatocytes. Direct evidence supporting this concept (11–14) has come from rodent experiments in which combined hepatocyte-islet clusters were placed in ectopic sites (21, 22). The present study has established that human hepatocytes are subject to the hepatotrophic influence of islet cell factors, and because dog islets were used with human hepatocytes in the nude mouse recipient, it could be shown that the hepatotrophic effect was not species-specific. These findings will be important in defining effective procedures for the maintenance of human hepatocytes both in vitro and in vivo, and for their use in hepatocellular transplantation, artificial liver, and gene therapy applications.

Previous studies have shown that mice can be protected from a lethal infection with the neurotropic JHM strain of mouse hepatitis virus (JHMV) by the adoptive transfer of delayed-type hypersensitivity (DTH)-inducer T cell clones specific for the virus. Protection does not involve the suppression of virus replication in the central nervous system (CNS) or via augmentation of the antiviral antibody response. In the present report we have compared the CNS lesions induced by JHMV in lethally infected and T cell clone protected mice. The presence of virus-specific T cell clones induced a transient increase in mononuclear cell infiltration into the parenchyma of the brains of protected mice, consistent with previous data suggesting that a DTH response was responsible for protection. Immunohistochemical studies suggested further that virus was not replicating in the ependyma or cellular infiltrate, but that the presence of the T cell clone prevented neuronal infection. While the mechanism of effectively altering the in vivo cellular tropism is unknown, survival is accompanied by increased specific destruction of target tissues with fulminant CNS demyelination and an increased incidence of persistent infection.

Bird flu, synonym of avian influenza (AI) caused by influenza A virus, become concern across the world for the possible incidence of the next human influenza pandemic. The latent danger of AI pandemic remains very real, though, the precise timing of occurrence and severity is uncertain. Each avian influenza type A (AIA) contains one of the 16 subtypes of haemagglutinin (HA) and 9 neuraminidases (NA) implicating theoretically 144 subtypes of AIA are possible in circulation, but only H1N1, H2N2 and H3N2 subtypes are documented for past pandemics in humans. In recent years H5N1, H7N3, H7N2, H7N7 and H9N2 are isolated from human samples, though H1N1 and H3N2 are still in circulation. Avian influenza viruses preferentially recognize receptor containing sialosugar chains terminating in sialic acid ?-2,3-galactose in bird, whereas, human preferentially contain ?-2,6-galactose subtype-receptor. To initiate a pandemic outbreak in human, the AIA viruses need alteration of receptor recognition specificity; and perfect match between HA and NA along with optimal cellular tropism. Cyclic nature of bird-flu emergence, and moreover, sporadic human incident reported around Asia and Europe in recent years anticipating a pandemic appearance of bird-flu in time to come. As we are on the edge of this alarming situation, AI prevention and containment can be considered under categories of surveillance, intervention, antiviral drugs, vaccination together with environment management issues.Keywords: Pandemic; Avian influenza; Genetic reassortment; Host specificity; Environment management DOI: http://dx.doi.org/10.3329/bjm.v25i1.4847Bangladesh J Microbiol, Volume 25, Number 1, June 2008, pp 1-8

Human herpesvirus 8 (HHV-8) or Kaposi's sarcoma associated herpesvirus (KSHV) is associated with Kaposi's sarcoma and primary effusion lymphoma. In vivo, HHV-8 DNA and transcripts have been detected in B cells, endothelial cells, macrophages, and epithelial cells. HHV-8 infects a variety of cell lines of human and animal origin, leading to latent or abortive infection. This study shows that the broad cellular tropism of HHV-8 may be in part due to its interaction with the ubiquitous host cell surface molecule, heparan sulfate (HS). This conclusion is based on the following findings: (i) HHV-8 infection of human foreskin fibroblast (HFF) cells was inhibited in a dose-dependent manner by soluble heparin, a glycosaminoglycan closely related to HS. Chondroitin sulfates A and C did not inhibit HHV-8 infection. (ii) Enzymatic removal of HFF cell surface HS with heparinase I and III reduced HHV-8 infection. (iii) Soluble heparin inhibited the binding of radiolabeled HHV-8 to human B cell lines, embryonic kidney epithelial (293) cells, and HFF cells, suggesting interference at the virus attachment stage. (iv) Cell surface adsorbed HHV-8 was displaced by soluble heparin. (v) Radiolabeled HHV-8 also bound to wild-type HS expressing Chinese hamster ovary (CHO-K1) cells. In contrast, binding of virus to mutant CHO cells deficient in HS was significantly reduced. These data show that the gamma2 herpesvirus HHV-8, similar to some members of alpha, beta, and gamma2 herpesviruses, adsorbs to cells by binding to cell surface HS-like moieties. Heparin did not completely prevent the binding and infectivity of HHV-8, suggesting that HHV-8 interactions with HS could be the first set of ligand-receptor interaction leading to the binding with one or more host cell receptors essential for the subsequent viral entry process.

Both a feral mouse ecotropic virus (WM-E) and Friend ecotropic virus (F-MuLV) were transmitted horizontally among adult mice. Infection resulted in the production of antiviral antibody in the recipients, with no evidence of viremia or clinical disease. However, persistent low-level virus replication was detectable in the spleens of these mice as long as 8 months after initial infection. External secretions, including saliva, semen, and uterine secretions from viremic mice contained high concentrations of infectious virus. Nevertheless, transmission occurred only from viremic males to either males or females. Male-to-male transmission appeared to occur by parenteral inoculation of infectious saliva during fighting behavior. Evidence is presented that infection of females was by the venereal route. Of four mouse strains examined, NFS/N, IRW, and C57L females were all susceptible to venereal infection, whereas AKR mice were not. Since AKR mice are susceptible to infection by WM-E administered parenterally, this resistance appeared to be mediated by local viral interference due to the high-level expression of endogenous Akv gp70 within the female reproductive tract. Although both WM-E and F-MuLV were transmitted from viremic males to females, infection by WM-E was significantly more efficient than that by F-MuLV. This difference correlated with a distinct difference in cellular tropism of WM-E and F-MuLV within the epididymis of viremic males. F-MuLV gp70 was expressed only within stromal elements, whereas WM-E gp70 was seen largely within the epithelial lining cells and luminal contents of the duct. No evidence of virus expression within germ cells was observed. The possible influence of virus expression by epithelial cells of the female reproductive tract on infection of embryos is discussed.

An understanding of the biologic characteristics and cellular tropism of human immunodeficiency virus (HIV) is critical to appreciate the diverse neurologic manifestations of HIV infection in patients with the acquired immunodeficiency syndrome (AIDS). Only carefully designed prospective studies can provide information regarding prevalence, incidence, and natural history of the full spectrum of neurologic complications of HIV infection. A degree of tropism for monocyte/macrophages and possibly for cells within the CNS seems certain. One of the most frequent complications is AIDS-related dementia, which reflects central nervous system invasion by HIV. Despite the evidence linking unchecked viral replication within the brain and progressive dementia, the basic pathogenetic mechanisms remain obscure. Further characterization of the cellular targets of HIV within the brain, and the mechanisms which ultimately lead to the dementia, is critical. The demonstration that HIV enters the central nervous system during the earliest stages of infection has major implications for antiviral agents which must penetrate brain parenchyma to clear the virus effectively. Other neurologic complications occur frequently, including myelopathies, peripheral neuropathies, opportunistic CNS infections, and CNS neoplasms. Many of these disorders are novel and incompletely characterized and their etiology is uncertain. While treatment is available for several of these conditions, it is generally not curative, and is often poorly tolerated because of adverse effects. Research directions will focus on better understanding of pathogenetic mechanisms, on earlier and more precise detection of these diverse conditions, and on improved therapeutic agents. For the future, efforts toward the development of a safe, effective vaccine are of critical importance. There are, however, already up to 2 million individuals in the United States who are already infected with HIV and who are thus at risk for developing 1 or several of these neurologic complications. Vaccination, even if it were available now, is not likely to benefit these individuals. While it is hoped that only a fraction of this infected population will develop neurologic symptoms, the prospects of an epidemic of AIDS-related dementia are ominous, particularly as antiviral therapy alone is unlikely to either eradicate the virus or restore brain function. In Africa and worldwide the numbers at risk for HIV-related diseases are enormous, and the risk factors for transmission of HIV less well defined. There, economic and medical resources are less than adequate to deal with a problem of this magnitude.

Several factors are involved in the high frequency of T-cell lymphomas of AKR mice, which appear mainly in the thymus at the age of 6–12 months [1]. The thymus is considered to play a major role in the disease since its removal prevents the development of T-cell lymphoma [2], while retransplantation of thymic epithelium to thymectomized AKR reconstitutes the high frequency of lymphoma [3], Although the AKR/J strain has the predisposition to develop the disease since birth [4], the mean latent period is delayed until the age of 8 months. The long latent period has been attributed to the delayed formation of the leukemogenic dual tropic virus (DTV) with the MCF characteristic [5, 6], formed in the thymus as a consequence of recombination within the envelope gene of ecotropic and xenotropic murine leukemia virus (MuLV). DTVs are detected only in preleukemic thymus and leukemic tissues of strains of mice prone to develop high incidence of leukemia [7]. DTVs enhance leukemia development whereas endogenous ecotropic or xenotropic viruses are usually nontumorigenic. Exceptional is the ecotropic virus isolate SL3 with the enhancing activity on T-cell lymphomagenesis [8]. These observations support the assumption that DTVs are proximal transforming agents of thymocytes and thereby responsible for high incidence of T lymphoma in AKR mice. Cloyd [9] proposed specific cellular tropism of two subclasses of MCF virus, and claimed that oncogenicity is closely linked to cellular differentiation. MCF isolated from lymphomatous thymus was replicating in the thymus and T peripheral cells, while nonlymphomagenic MCF isolated from leukemic spleen of NFS mice did not replicate in the thymus but rather in bone marrow cells, spleen, and lymph node B-lymphocytes.

The biologic, serologic, and molecular properties of isolates of human immunodeficiency virus type 1 (HIV-1) from the central nervous system (CNS) were determined and compared to those of isolates from peripheral blood and lymph nodes. Among these were pairs of CNS and blood isolates obtained from six infected individuals. The data show that HIV-1 isolates from the CNS can be distinguished from peripheral blood isolates by their (i) relative inability to infect established T-cell lines, (ii) reduced cytopathogenicity, (iii) inability to modulate CD4 antigen expression on infected cells, (iv) efficient replication in peripheral blood macrophages, and (v) insensitivity to serum neutralization. Paired CNS and peripheral blood isolates from the same individual also display some differences in cellular tropism. The blood isolates replicate better in T-cell lines and glioma cell lines, whereas the paired CNS isolates replicate more efficiently in primary macrophages. These results suggest that viruses isolated from the CNS of infected individuals may represent a specific HIV-1 subgroup.

Strains of the murine coronavirus mouse hepatitis virus type 4 (MHV-4) which contained a mutation in the E2 peplomer glycoprotein were obtained by selection for resistance to neutralization by monoclonal antibodies. Characterization of six variants representing two independent epitopes on E2, E2B and E2C, by in vitro neutralization and antibody-binding assays demonstrated that selection for an alteration in epitope E2B also resulted in changes in epitope E2C and vice versa. We observed a mutation frequency of approximately 10(-4.3) to 10(-4.6), which is consistent with the expected occurrence of single point mutations. The variant virus strains were attenuated with respect to neurovirulence when compared with wild-type MHV-4. Mice normally develop encephalomyelitis and die after wild-type MHV-4 infection. Mice receiving 2- to 3-log-higher doses of the variant strains survived and developed demyelinating disease. As the disease progressed, evidence of remyelination and ongoing demyelination was observed up to 65 days after infection. Virus reisolated 15 days after infection retained the variant phenotype. The data indicate that the E2 glycoprotein plays a central role in determining the cellular tropism and virulence of MHV-4 in the mouse.

Summary Nude (nu/nu) mice were inoculated intracranially with 106.5 newborn mouse 50% lethal doses of K virus and were studied over a period of 28 weeks using serological methods, virus assay and immunohistological staining for viral antigens. K virus infection of nude mice, although clinically asymptomatic, was slowly progressive despite prompt IgM and IgG antibody response. The highest titres of K virus infectivity were reached in spleens, kidneys and intestines. Vascular endothelial cells represented the major site of viral replication, as has been shown to be the case in immunologically normal mice, with extensive involvement of intestinal capillaries. In addition, however, unlike immunologically normal mice, nude mice inoculated with K virus developed multifocal infection of renal tubular epithelial cells. Nude mice did not develop histologically detectable evidence of central nervous system involvement by K virus, and K virus infection did not result in neoplasia. Infected vascular endothelial cells and renal tubular epithelial cells in animals studied at 16 and 27 weeks after inoculation were grouped in scattered clusters, suggesting local spread of infection. The present study indicates that nude mice with preserved B cell function but impaired T cell-mediated immunity are able to limit systemic dissemination of K virus but are unable to prevent local progression of infection by cell-to-cell spread. K virus is capable of altering its cellular tropism during chronic infection.

Evolution of virus expression in various lymphoid tissues of C 3 H/Fe mice, inoculated with 2 variants of tissue culture adapted Gross Passage-A virus, was studied with the aid of parallel in vitro XC co-culture technique and electron microscopic examination. When the mice were neonatally inoculated with a highly leukemogenic variant (TGV virus), 3 phases of virus expression could be distinguished during the average 2 months of latent period. In the early phase (for the first 3 weeks after virus inoculation) as well as in the later phase (after the 50 th day ), the virus could be detected abundantly in bone marrows and spleens, moderately in thymuses and slightly or not at all in lymph nodes and kidneys. In the intermediate phase (the 20 th to the 50 th days ), the virus disappeared completely or decreased significantly from all tissues tested. In the case of mice similarly inoculated with a non-leukemogenic variant (N 1 virus), no virus could be detected in all tissues during the 2 months period after virus inoculation. The virus recovered from in vitro explanted and cultured kidney cells, taken from mice inoculated neonatally with TGV virus, induced typical Gross type lymphoid leukemia in all C 3 HeB/Fe mice inoculated as newborns. However, in vitro cellular tropism of this virus was revealed as B tropic while that of the original TGV virus was N tropic. Frequent differentiation to heterologous tissues was observed by electron microscope in the thymuses of mice inoculated as newborns with TGV virus.

The cellular tropism of hepatitis C virus (HCV) was studied in vivo in samples from patients with persistent HCV infection. Plasma, liver, peripheral blood mononuclear cell (PBMC), and bone marrow cell (BMC) samples from 15 subjects positive for anti-HCV antibodies were tested for the presence of HCV RNA sequences by reverse transcription PCR. Virus-specific RNA sequences were found to be present in liver samples from all subjects (100%), in plasma samples from 13 of 15 patients (86.7%), in PBMC samples from 3 patients (20%), and in BMC samples from 9 (60%) of the 15 anti-HCV-positive patients enrolled in this study. The presence of the molecular intermediate of HCV replication (the negative-stranded HCV RNA) was evident in the two of the three PBMC and in five of the nine BMC HCV RNA-positive samples. Finally, we studied the nucleotide sequence of a large portion (-270 to -59) of the 5'untranslated region of HCV amplified from plasma samples of 12 of the 15 patients with and without HCV in BMCs; the degree of heterogeneity compared with the prototype HCV sequence was similar in both groups. The data principally indicate that HCV infection of PBMCs and BMCs is frequent in persistently infected patients, as shown by the occurrence of positive- and negative-stranded HCV RNA, thus suggesting the possibility of extrahepatic replication of HCV.

Background Measles virus attenuation has been historically performed by adaptation to cell culture. The current dogma is that attenuated virus strains induce more type I IFN and are more resistant to IFN-induced protection than wild type (wt). Results The adaptation of a measles virus isolate (G954-PBL) by 13 passages in Vero cells induced a strong attenuation of this strain in vivo. The adapted virus (G954-V13) differs from its parental strain by only 5 amino acids (4 in P/V/C and 1 in the M gene). While a vaccine strain, Edmonston Zagreb, could replicate equally well in various primate cells, both G954 strains exhibited restriction to the specific cell type used initially for their propagation. Surprisingly, we observed that both G954 strains induced type I IFN, the wt strain inducing even more than the attenuated ones, particularly in human plasmacytoid Dendritic Cells. Type I IFN-induced protection from the infection of both G954 strains depended on the cell type analyzed, being less efficient in the cells used to grow the viral strain. Conclusion Thus, mutations in M and P/V/C proteins can critically affect MV pathogenicity, cellular tropism and lead to virus attenuation without interfering with the α/β IFN system.